Effectiveness and Functional Mechanism of a Multicomponent Sanitizer against Biofilms Formed by Escherichia coli O157:H7 and Five Salmonella Serotypes Prevalent in the Meat Industry

2020 ◽  
Vol 83 (4) ◽  
pp. 568-575
Author(s):  
RONG WANG ◽  
YOU ZHOU ◽  
NORASAK KALCHAYANAND ◽  
DAYNA M. HARHAY ◽  
TOMMY L. WHEELER

ABSTRACT Biofilm formation by Escherichia coli O157:H7 and Salmonella enterica at meat processing plants poses a potential risk of meat product contamination. Many common sanitizers are unable to completely eradicate biofilms formed by these foodborne pathogens because of the three-dimensional biofilm structure and the presence of bacterial extracellular polymeric substances (EPSs). A novel multifaceted approach combining multiple chemical reagents with various functional mechanisms was used to enhance the effectiveness of biofilm control. We tested a multicomponent sanitizer consisting of a quaternary ammonium compound (QAC), hydrogen peroxide, and the accelerator diacetin for its effectiveness in inactivating and removing Escherichia coli O157:H7 and Salmonella enterica biofilms under meat processing conditions. E. coli O157:H7 and Salmonella biofilms on common contact surfaces were treated with 10, 20, or 100% concentrations of the multicomponent sanitizer solution for 10 min, 1 h, or 6 h, and log reductions in biofilm mass were measured. Scanning electron microscopy (SEM) was used to directly observe the effect of sanitizer treatment on biofilm removal and bacterial morphology. After treatment with the multicomponent sanitizer, viable E. coli O157:H7 and Salmonella biofilm cells were below the limit of detection, and the prevalence of both pathogens was low. After treatment with a QAC-based control sanitizer, surviving bacterial cells were countable, and pathogen prevalence was higher. SEM analysis of water-treated control samples revealed the three-dimensional biofilm structure with a strong EPS matrix connecting bacteria and the contact surface. Treatment with 20% multicomponent sanitizer for 10 min significantly reduced biofilm mass and weakened the EPS connection. The majority of the bacterial cells had altered morphology and compromised membrane integrity. Treatment with 100% multicomponent sanitizer for 10 min dissolved the EPS matrix, and no intact biofilm structure was observed; instead, scattered clusters of bacterial aggregates were detected, indicating the loss of cell viability and biofilm removal. These results indicate that the multicomponent sanitizer is effective, even after short exposure with dilute concentrations, against E. coli O157:H7 and S. enterica biofilms. HIGHLIGHTS

2018 ◽  
Vol 19 (0) ◽  
Author(s):  
Priscila Alves Dias ◽  
Daiani Teixeira Silva ◽  
Cláudio Dias Timm

Resumo Kefir é o produto da fermentação do leite pelos grãos de kefir. Esses grãos contêm uma mistura simbiótica de bactérias e leveduras imersas em uma matriz composta de polissacarídeos e proteínas. Muitos benefícios à saúde humana têm sido atribuídos ao kefir, incluindo atividade antimicrobiana contra bactérias Gram positivas e Gram negativas. A atividade antimicrobiana de 60 microrganismos isolados de grãos de kefir, frente à Escherichia coli O157:H7, Salmonella enterica subsp. enterica sorotipos Typhimurium e Enteritidis, Staphylococcus aureus e Listeria monocytogenes, foi estudada através do teste do antagonismo. A ação antimicrobiana dos sobrenadantes das bactérias ácido-lácticas que apresentaram atividade no teste do antagonismo foi testada. O experimento foi repetido usando sobrenadantes com pH neutralizado. Salmonella Typhimurium e Enteritidis sobreviveram por 24 horas no kefir em fermentação. E. coli O157:H7, S. aureus e L. monocytogenes foram recuperados até 72 horas após o início da fermentação. Todos os isolados apresentaram atividade antimicrobiana contra pelo menos um dos patógenos usados no teste do antagonismo. Sobrenadantes de 25 isolados apresentaram atividade inibitória e três mantiveram essa atividade com pH neutralizado. As bactérias patogênicas estudadas sobreviveram por tempo superior àquele normalmente utilizado para a fermentação do kefir artesanal, o que caracteriza perigo em potencial para o consumidor quando a matéria-prima não apresentar segurança sanitária. Lactobacillus isolados de grãos de kefir apresentam atividade antimicrobiana contra cepas de E. coli O157:H7, Salmonella sorotipos Typhimurium e Enteritidis, S. aureus e L. monocytogenes além daquela exercida pela diminuição do pH.


2006 ◽  
Vol 69 (8) ◽  
pp. 1978-1982 ◽  
Author(s):  
J. E. MANN ◽  
M. M. BRASHEARS

In order to provide beef processors with valuable data to validate critical limits set for temperature during grinding, a study was conducted to determine Escherichia coli O157:H7 growth at various temperatures in raw ground beef. Fresh ground beef samples were inoculated with a cocktail mixture of streptomycin-resistant E. coli O157:H7 to facilitate recovery in the presence of background flora. Samples were held at 4.4, 7.2, and 10°C, and at room temperature (22.2 to 23.3°C) to mimic typical processing and holding temperatures observed in meat processing environments. E. coli O157:H7 counts were determined by direct plating onto tryptic soy agar with streptomycin (1,000 μg/ml), at 2-h intervals over 12 h for samples held at room temperature. Samples held under refrigeration temperatures were sampled at 4, 8, 12, 24, 48, and 72 h. Less than one log of E. coli O157:H7 growth was observed at 48 h for samples held at 10°C. Samples held at 4.4 and 7.2°C showed less than one log of E. coli O157:H7 growth at 72 h. Samples held at room temperature showed no significant increase in E. coli O157:H7 counts for the first 6 h, but increased significantly afterwards. These results illustrate that meat processors can utilize a variety of time and temperature combinations as critical limits in their hazard analysis critical control point plans to minimize E. coli O157:H7 growth during the production and storage of ground beef.


2002 ◽  
Vol 65 (8) ◽  
pp. 1215-1220 ◽  
Author(s):  
CHIA-MIN LIN ◽  
SARAH S. MOON ◽  
MICHAEL P. DOYLE ◽  
KAY H. McWATTERS

Iceberg lettuce is a major component in vegetable salad and has been associated with many outbreaks of foodborne illnesses. In this study, several combinations of lactic acid and hydrogen peroxide were tested to obtain effective antibacterial activity without adverse effects on sensory characteristics. A five-strain mixture of Escherichia coli O157:H7, Salmonella enterica serotype Enteritidis, and Listeria monocytogenes was inoculated separately onto fresh-cut lettuce leaves, which were later treated with 1.5% lactic acid plus 1.5% hydrogen peroxide (H2O2) at 40°C for 15 min, 1.5% lactic acid plus 2% H2O2 at 22°C for 5 min, and 2% H2O2 at 50°C for 60 or 90 s. Control lettuce leaves were treated with deionized water under the same conditions. A 4-log reduction was obtained for lettuce treated with the combinations of lactic acid and H2O2 for E. coli O157:H7 and Salmonella Enteritidis, and a 3-log reduction was obtained for L. monocytogenes. However, the sensory characteristics of lettuce were compromised by these treatments. The treatment of lettuce leaves with 2% H2O2 at 50°C was effective not only in reducing pathogenic bacteria but also in maintaining good sensory quality for up to 15 days. A ≤4-log reduction of E. coli O157:H7 and Salmonella Enteritidis was achieved with the 2% H2O2 treatment, whereas a 3-log reduction of L. monocytogenes was obtained. There was no significant difference (P > 0.05) between pathogen population reductions obtained with 2% H2O2 with 60- and 90-s exposure times. Hydrogen peroxide residue was undetectable (the minimum level of sensitivity was 2 ppm) on lettuce surfaces after the treated lettuce was rinsed with cold water and centrifuged with a salad spinner. Hence, the treatment of lettuce with 2% H2O2 at 50°C for 60 s is effective in initially reducing substantial populations of foodborne pathogens and maintaining high product quality.


2007 ◽  
Vol 70 (4) ◽  
pp. 841-850 ◽  
Author(s):  
JOSH R. BRANEN ◽  
MARTHA J. HASS ◽  
ERIN R. DOUTHIT ◽  
WUSI C. MAKI ◽  
A. LARRY BRANEN

Enzymatic bio-nanotransduction is a biological detection scheme based on the production of nucleic acid nano-signals (RNA) in response to specific biological recognition events. In this study, we applied an enzymatic bio-nanotransduction system to the detection of important food-related pathogens and a toxin. Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium, and staphylococcal enterotoxin B (SEB) were chosen because of the implications of these targets to food safety. Primary antibodies to each of the targets were used to functionalize magnetic beads and produce biological recognition elements (antibodies) conjugated to nano-signal–producing DNA templates. Immunomagnetic capture that was followed by in vitro transcription of DNA templates bound to target molecules produced RNA nano-signals specific for every target in the sample. Discrimination of RNA nano-signals with a standard enzyme-linked oligonucleotide fluorescence assay provided a correlation between nano-signal profiles and target concentrations. The estimated limit of detection was 2.4 × 103 CFU/ml for E. coli O157:H7, 1.9 × 104 CFU/ml for S. enterica serovar Typhimurium, and 0.11 ng/ml for SEB with multianalyte detection in buffer. Low levels of one target were also detected in the presence of interference from high levels of the other targets. Finally, targets were detected in milk, and detection was improved for E. coli O157 by heat treatment of the milk.


2006 ◽  
Vol 50 (8) ◽  
pp. 2789-2796 ◽  
Author(s):  
Lucas D. Tilley ◽  
Orion S. Hine ◽  
Jill A. Kellogg ◽  
Jed N. Hassinger ◽  
Dwight D. Weller ◽  
...  

ABSTRACT The objective was to improve efficacy of antisense phosphorodiamidate morpholino oligomers (PMOs) by improving their uptake into bacterial cells. Four different bacterium-permeating peptides, RFFRFFRFFXB, RTRTRFLRRTXB, RXXRXXRXXB, and KFFKFFKFFKXB (X is 6-aminohexanoic acid and B isβ -alanine), were separately coupled to two different PMOs that are complementary to regions near the start codons of a luciferase reporter gene (luc) and a gene required for viability (acpP). Luc peptide-PMOs targeted to luc inhibited luciferase activity 23 to 80% in growing cultures of Escherichia coli. In cell-free translation reactions, Luc RTRTRFLRRTXB-PMO inhibited luciferase synthesis significantly more than the other Luc peptide-PMOs or the Luc PMO not coupled to peptide. AcpP peptide-PMOs targeted to acpP inhibited growth of E. coli or Salmonella enterica serovar Typhimurium to various extents, depending on the strain. The concentrations of AcpP RFFRFFRFFXB-PMO, AcpP RTRTRFLRRTXB-PMO, AcpP KFFKFFKFFKXB-PMO, and ampicillin that reduced CFU/ml by 50% after 8 h of growth (50% inhibitory concentration [IC50]) were 3.6, 10.8, 9.5, and 7.5μ M, respectively, in E. coli W3110. Sequence-specific effects of AcpP peptide-PMOs were shown by rescuing growth of a merodiploid strain that expressed acpP with silent mutations in the region targeted by AcpP peptide-PMO. In Caco-2 cultures infected with enteropathogenic E. coli (EPEC), 10 μM AcpP RTRTRFLRRTXB-PMO or AcpP RFFRFFRFFXB-PMO essentially cleared the infection. The IC50 of either AcpP RTRTRFLRRTXB-PMO or AcpP RFFRFFRFFXB-PMO in EPEC-infected Caco-2 culture was 3 μM. In summary, RFFRFFRFFXB, RTRTRFLRRTXB, or KFFKFFKFFXB, when covalently bonded to PMO, significantly increased inhibition of expression of targeted genes compared to PMOs without attached peptide.


1997 ◽  
Vol 60 (2) ◽  
pp. 102-106 ◽  
Author(s):  
LAURA CABEDO ◽  
JOHN N. SOFOS ◽  
GLENN R. SCHMIDT ◽  
GARY C. SMITH

Three strains of Escherichia coli O157:H7 were grown in tryptic soy broth (TSB) or in a sterile cattle manure extract at 35°C for 18 ± 2 h. Aliquots from both inocula containing 106 CFU/ml were used to inoculate 1-cm3 cubes of beef muscle or adipose tissue by immersion for 20 min at 21°C. After removal from the inoculum, one-half of the samples were analyzed for bacterial cell numbers and pH, and the other half were stored at 4°C for 2 or 3 h before analysis. Samples were analyzed by enumerating bacteria present in liquid droplets deposited on the tissue and bacteria loosely or strongly attached to the tissue in order to determine attachment strength. Total numbers of cells on beef muscle tissue (bacteria in liquid droplets, as well as those loosely and strongly attached) were 5.65 ± 0.14 and 5.76 ± 0.26 log CFU/cm2 for E. coli O157:H7 inocula grown in TSB and manure extract, respectively. The differences in attachment strength between inocula from the two media were not significant (P > 0.05). A 2-h storage period after exposure of muscle tissue to an E. coli O157:H7 inoculum did not influence attachment strength. Numbers of bacteria attached to adipose tissue and muscle (5.31 ± 0.08 and 5.48 ± 0.09 log CFU/cm2, respectively) were not significantly different (P > 0.05). After 3 h at 4°C, the attachment strength of E. coli O157:H7 cells on muscle or adipose tissue had not changed. Overall, the culture medium and type of beef tissue did not affect the numbers of E. coli O157:H7 cells attached, nor the strength of their attachment, to muscle or adipose tissue.


2013 ◽  
Vol 76 (7) ◽  
pp. 1245-1249 ◽  
Author(s):  
F. BREIDT ◽  
K. KAY ◽  
J. COOK ◽  
J. OSBORNE ◽  
B. INGHAM ◽  
...  

A critical factor in ensuring the safety of acidified foods is the establishment of a thermal process that assures the destruction of acid-resistant vegetative pathogenic and spoilage bacteria. For acidified foods such as dressings and mayonnaises with pH values of 3.5 or higher, the high water phase acidity (acetic acid of 1.5 to 2.5% or higher) can contribute to lethality, but there is a lack of data showing how the use of common ingredients such as acetic acid and preservatives, alone or in combination, can result in a 5-log reduction for strains of Escherichia coli O157:H7, Salmonella enterica, and Listeria monocytogenes in the absence of a postpackaging pasteurization step. In this study, we determined the times needed at 10°C to achieve a 5-log reduction of E. coli O157:H7, S. enterica, and L. monocytogenes in pickling brines with a variety of acetic and benzoic acid combinations at pH 3.5 and 3.8. Evaluation of 15 different acid-pH combinations confirmed that strains of E. coli O157:H7 were significantly more acid resistant than strains of S. enterica and L. monocytogenes. Among the acid conditions tested, holding times of 4 days or less could achieve a 5-log reduction for vegetative pathogens at pH 3.5 with 2.5% acetic acid or at pH 3.8 with 2.5% acetic acid containing 0.1% benzoic acid. These data indicate the efficacy of benzoic acid for reducing the time necessary to achieve a 5-log reduction in target pathogens and may be useful for supporting process filings and the determination of critical controls for the manufacture of acidified foods.


2013 ◽  
Vol 76 (4) ◽  
pp. 668-673 ◽  
Author(s):  
CHRIS TIMMONS ◽  
SHEFALI DOBHAL ◽  
JACQUELINE FLETCHER ◽  
LI MARIA MA

Foodborne illnesses caused by Salmonella enterica and Escherichia coli O157:H7 are worldwide health concerns. Rapid, sensitive, and robust detection of these pathogens in foods and in clinical and environmental samples is essential for routine food quality testing, effective surveillance, and outbreak investigations. The aim of this study was to evaluate the effect on PCR sensitivity of adding a short, AT-rich overhanging nucleotide sequence (flap) to the 5′ end of PCR primers specific for the detection of Salmonella and E. coli O157:H7. Primers targeting the invA gene of Salmonella and the rfbE gene of E. coli O157:H7 were synthesized with or without a 12-bp, AT-rich 5′ flap (5′-AATAAATCATAA-3′). Singleplex PCR, multiplex PCR, and real-time PCR sensitivity assays were conducted using purified bacterial genomic DNA and crude cell lysates of bacterial cells. The effect of background flora on detection was evaluated by spiking tomato and jalapeno pepper surface washes with E. coli O157:H7 and Salmonella Saintpaul. When targeting individual pathogens, end-point PCR assays using flap-amended primers were more efficient than nonamended primers, with 20.4 and 23.5% increases in amplicon yield for Salmonella and E. coli O157:H7, respectively. In multiplex PCR assays, a 10- to 100-fold increase in detection sensitivity was observed when the primer flap sequence was incorporated. This improvement in both singleplex and multiplex PCR efficiency and sensitivity can lead to improved Salmonella and E. coli O157:H7 detection.


2005 ◽  
Vol 68 (11) ◽  
pp. 2443-2446 ◽  
Author(s):  
ISABEL C. BLACKMAN ◽  
YOUNG W. PARK ◽  
MARK A. HARRISON

An oxidative complex composed of ferric iron chloride hexahydrate, ADP, and ascorbic acid can generate hydrogen peroxide and hydroxyl radicals in fibroblasts. These compounds are naturally found in meat and meat-based products and may elicit oxidative stress on Escherichia coli O157:H7, thus conferring thermotolerance to the bacterium due to the phenomenon of the global stress response. The effect of the levels of the oxidative complex on the thermotolerance of E. coli O157:H7 was investigated. Cultures of E. coli O157:H7 strains EO139 and 380-94 were mixed in three different concentrations (10:10: 40, 15:15:60, and 20:20:80 μM) of the oxidative complex (iron III chloride, ADP, and ascorbic acid, respectively). The samples were inserted into capillary tubes and heated in a circulating water bath at 59 and 60°C for EO139 and 380-94, respectively. Tubes were removed at intervals of 5 min for up to 1 h and contents spirally plated on plate count agar that was incubated for 48 h at 37°C. The thermotolerance of both E. coli O157:H7 strains EO139 and 380-94 was influenced by the concentrations of the oxidative complex. The ratio of 10:10:40 μM enhanced thermotolerance of EO139 and 390-94 at 59 and 60°C, respectively. However, exposure to the ratios of 15:15:60 and 20:20:80 μM rendered the pathogen more sensitive to the lethal effect and did not enhance the thermotolerance of the cells. The significance of this study is twofold. This experiment proves that oxidative stress can enhance thermotolerance of bacterial cells induced by an oxidative complex if only in a specific ratio and concentration. It is possible to speculate that if the chemical compounds are present in this ratio in meats, they may enhance the thermal resistance of E. coli O157:H7 and make the bacteria more difficult to eliminate, thus increasing the risk of foodborne illness in consumers.


2012 ◽  
Vol 75 (12) ◽  
pp. 2213-2218 ◽  
Author(s):  
JOHN W. SCHMIDT ◽  
TERRANCE M. ARTHUR ◽  
JOSEPH M. BOSILEVAC ◽  
NORASAK KALCHAYANAND ◽  
TOMMY L. WHEELER

Bacteria are known to be present in the air at beef processing plants, but published data regarding the prevalences of airborne Escherichia coli O157:H7 and Salmonella enterica are very limited. To determine if airborne pathogens were present in beef processing facilities, we placed sedimentation sponges at various locations in three commercial beef plants that processed cattle from slaughter through fabrication. For the 291 slaughter area air samples, E. coli O157:H7 was isolated from 15.8% and S. enterica from 16.5%. Of the 113 evisceration area air samples, E. coli O157:H7 was isolated from only one sample and S. enterica was not isolated from any sample. Pathogens were not isolated from any of the 87 air samples from fabrication areas. Pathogen prevalences, aerobic plate counts, and Enterobacteriaceae counts were highest for air samples obtained from locations near hide removal operations. The process of hide removal disperses liquid droplets, which may contact neighboring carcasses. Samples were obtained both from hide removal locations that were close enough to hide pullers to be contacted by droplets and from locations that were not contacted by droplets. Higher pathogen prevalences, aerobic plate counts, and Enterobacteriaceae counts were observed at locations with samples contacted by the hide removal droplets. We conclude that the hide removal processes likely introduce pathogens into the air via a dispersion of liquid droplets and that these droplets may be an underappreciated source of hide-to-carcass contamination.


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